This proposal is specifically designed to define a lead vaginal topical microbicide compound from a series of 78 pyrimidinedione molecules synthesized by the Samjin Pharmaceutical Co., Ltd. Topical microbicides represent an important strategy for preventing the transmission of HIV through sexual intercourse, the predominant mode by which HIV is transmitted worldwide. The specific rationale for the development of the pyrimidinedione series of molecules includes the observation that the compounds are active against both HIV-1 and HIV-2, the compounds inhibit HIV through two distinct mechanisms of action, including inhibition of reverse transcription and viral entry, and display a significantly high in vitro therapeutic index (0.5 to 4 million) against HIV. The specific aims of the proposal are to define the comparative topical microbicide efficacy and toxicity of the test molecules and to define lead microbicidal candidates for further evaluation. The characteristics to be used to select for superior microbicides will include efficacy in virus entry inhibition assays, against clinical strains of HIV-1 and HIV-2 in PBMCs, and in cell-free and cell-associated virus transmission assays, as well as toxicity to Lactobacilli and normal human cells. These assays are designed to address the FDA's points to consider in the preclinical development of anti-infectives, and are specifically tailored to address the requirements of an effective topical microbicide. In light of the broad spectrum of antiviral properties of the pyrimidinedione series with regard to the ability of distinct compounds to inhibit virus entry and reverse transcription, we propose to define the ability of a combination of pyrimidinedione molecules to be used as a component of a microbicide cocktail therapy with or without other approved anti- HIV agents or microbicide candidates. In addition we propose to validate the in vitro assay results in an established ex vivo tissue culture model of HIV infection and drug efficacy using the MatTek cervical explant tissue model. The deveopment of the results of this research will be used as a basis for the submission of a Phase II SBIR proposal that will further the preclincal development towards commercialization of the lead compound(s). Goals of the Phase II proposal will be optimization of compound formulation, continuation of preclinical trials with relevent animal models of infection to examine compound bioavailability, efficacy, toxicity, and pharmocokinetics in preparation for clinical therapeutic assessments. [unreadable] [unreadable]